Chemical Crosslinking of Myosin Subfragment-1 to F-Actin in the Presence of Nucleotide1

F-Actin and myosin subfragment-1 (S-l) were covalently crosslinked in the absence and presence of nucleotides, adenyl-5'-yl imidodiphosphate (AMPPNP), ATP, and ADP, at various KC1 concentrations (0-0.5 M), using a zero-length crosslinker, 1-ethyl-3-[3-(dimethylamino)propyl]carbodiimide (EDC). The rate of production of the covalent acto-S-1 complex was almost proportional to the amount of the acto-S-l (-nucleotide) complex existing in the reaction medium. However, the Mg-ATPase activity of the covalent acto-S-1 complex thus produced was affected by the presence of nucleotides. When S-l was crosslinked to F-actin in the absence of nucleotide at 0-0.5 M KC1, the Mg-ATPase activity of S-l was enhanced from 0.1-0.3 to 12-15 s"1., The Mg-ATPase activity of covalent complex produced in the presence of AMPPNP at 0 M KC1 was 13 s-1 which was equal to that produced in the absence of nucleotide. The activity decreased with increasing KC1 concen-tration of the crosslinking medium, reaching 6 s-1 at 0.5 M KC1. Covalent acto-S-1 complex was also produced when acto-S-1-ADP-P was formed during ATP hydrol-ysis. The Mg-ATPase activity of the covalent acto-S-1 complex crosslinked durin